Shortly after the introduction of antibiotics in the 1940's, strains of bacteria that were resistant to these drugs began to emerge. Today, the continued emergence of bacterial strains that are resistant to multiple broad classes of antibiotics has raised significant concern among the general medical community, some even suggesting we may be about to enter a post-antibiotic era. Among resistant bacteria, methicillin-resistant Staphylococcus aureus (MRSA) is an agent of major concern. Bacteriophages, viruses that infect bacteria, have been proposed as biological control agents for the treatment of bacterial infections in humans and animals. Phage therapy against S. aureus and MRSA is facilitated by their broad susceptibility to a single type of phages, referred to as the Twort-like group. While there is considerable interest in the use of phages as therapeutics, an important but currently underdeveloped field of research is the biology of the phages themselves. This proposal is divided into two major aims: the first is to study the genetics and biology of the Twort-like phage K, and the second is to leverage this study to develop methods that can be used to engineer or further study the biology of this phage. In the first aim, the library of suppressible mutants will be generated, providing a map of most of the essential genes in this phage and allowing functions to be assigned to a number of unknown genes. The second aim of the project is to develop new methods for genetic manipulation of phages, primarily by developing a selectable marker system that can be used in the phage genetic context. This system will allow phage genes to be inactivated or replaced at will, allowing a number of future fundamental studies to be conducted. The strategies developed here should also be useful in other phage systems, thus catalyzing progress in the field of phage biology as a whole.